1. Field of the Invention
The present invention relates generally to medicines and pharmaceutical products derived from plants, and specifically to a process of extracting TAXOL.RTM. from the leaves of taxus cuspidata plants.
2. Description of the Prior Art
TAXOL.RTM. (paclitaxel) is one of a class of drugs called taxanes. These types of drugs promote polymerization of tubulin and stabilize the structure of intracellular microtubules. This process has the effect of inhibiting the normal dynamic reorganization of the microtubules that is necessary for interphase and mitotic functions. Paclitaxel may also potentiate the cytotoxic effects of radiation.
The pharmacokinetics of paclitaxel vary considerably depending on the dosage and duration of infusion. Plasma concentrations of paclitaxel decline in a biphasic manner following injection. Increasing a 24-hour infusion from 135 mg/mL to 175 mg/mL increased Cmax by 87% while the area under the plasma concentration-time curve (AUC) remained constant. Increasing a 3-hour infusion increased Cmax by 68% and the AUC by 89%. With the 24-hour infusion of paclitaxel, the mean apparent volume of distribution in the steady state ranged from 227 to 688 L/m2, indicating extensive extravascular distribution or tissue binding. In vitro studies show that 89% to 98% of paclitaxel is bound to human serum proteins. Paclitaxel may undergo hepatic metabolism in humans. Studies have demonstrated extensive nonrenal clearance. The disposition of paclitaxel in patients with renal or hepatic dysfunction has not yet been determined.
In two studies of ovarian cancer patients receiving paclitaxel in doses of 135 mg/mL or 175 mg/mL, the response rates were 22% and 30%, respectively, with 6 complete and 18 partial responses among 92 patients. Duration of response was 7.2 and 7.5 months, respectively, with median survival rates of 8.1 months and 15.9 months. A Phase III study of 471 breast cancer patients receiving paclitaxel in doses of 135 mg/mL or 175 mg/mL infused over 3 hours indicated an overall response rate of 26%, with 17 complete and 99 partial responses. The median duration of response was 3.5 months, and the median survival time was 11.7 months.
TAXOL.RTM. should not be used for patients with hypersensitivity to polyoxyethylated castor oil (Cremaphor EL) or TAXOL.RTM.. Patients should be pretreated with corticosroids (e.g., dexamethasone and diphenhydramine) and H2-receptor antagonists (cimetidine or ranitidine.) Patients receiving concomitant ketaconazole should be treated with caution. Two percent of patients receiving Taxol have severe hypersensitivity reactions that are characterized by dyspnea, hypotension, angioedema, and generalized urticaria. One incident of progressive hypotension, possibly related to the use of TAXOL.RTM., resulted in the death of the patient. Other side effects include bone marrow suppression, abnormal ECG, peripheral neuropathy, myalgia and arthralgia, alopecia, injection site reaction, nausea, vomiting, diarrhea, and mucositis. Continuing safety surveillance has produced rare reports of hepatic necrosis, hepatic encephalopathy, phlebitis, and cellulitis. Available as a clear, colorless to slightly yellow, viscous solution, TAXOL.RTM. is packaged in single-dose vials containing 30 mg/5 mL. After dilution with a suitable parenteral fluid to 0.3 to 1.2 mg/mL, 135 mg/m2 paclitaxel is infused intravenously over 24 hours every 3 weeks for ovarian cancer. The dose level for breast cancer is 175 mg/mL, administered in the same manner. TAXOL.RTM. treatment should not be repeated until neutrophil and platelet counts return to 1500 cells/mm2 and 100,000 cells/mm2, respectively.
The National Cancer Institute performed research on TAXOL.RTM. in the 1960's, examining some 35,000 botanical species in search for plants that could yield cancer-inhibiting products. The cancer-inhibiting material extracted from the bark of Taxus Brevifolia was reported to have beneficial effects upon patients suffering from leukemia, lung cancer, and other types of cancer. According to statistics issued by the National Cancer Institute, TAXOL.RTM. is reported to have cured 30% of the patients suffering from ovarian cancer who received dosages of TAXOL.RTM.. Likewise, 50% of patients suffering from breast cancer and 20% of patients suffering from lung cancer also were cured by the application of TAXOL.RTM.. Schiff additionally reported the effectiveness of TAXOL.RTM. in impeding the growth of cancers. (Schiff, P. B., J. Fant and S. B. Horwitz, Nature, 277, 665 (1979)) In addition, the analytical analysis of TAXOL.RTM. was accomplished in 1971. (E. K. Rowinsky et al., J. National Can. Inst., 82, 11247 (1990))
In December of 1992, the Food and Drug Administration (FDA) approved the natural form of paclitaxel for treatment of metastatic ovarian cancer after failure of first-line or subsequent chemotherapy, and the use of paclitaxel for the treatment of metastatic breast cancer received marketing approval in April of 1994.
TAXOL.RTM. is a complex compound containing unsaturated carbons. Although researchers reported total synthesis of TAXOL.RTM. in 1994, the process was so expensive that it was deemed that it was not commercially feasible to produce this form of TAXOL.RTM. on a production level. Therefore, TAXOL.RTM. can only be obtained currently through natural sources, namely yew trees.
Accordingly, the demand for TAXOL.RTM. is ever increasing. The supply of TAXOL.RTM., however, is not sufficient to provide for the needs of all the patients who are currently suffering from cancer. TAXOL.RTM. is so scarce that adequate amounts cannot be provided for the research of testing its effectiveness against skin cancer and other types of lung cancer.
Presently, TAXOL.RTM. is primarily obtained from the bark of yew trees. This bark contains about 0.02% TAXOL by weight. Because the bark must be removed from the tree in order to effectively extract the TAXOL.RTM., the tree subsequently dies after the removal of its bark. This restricts the amount of TAXOL.RTM. which can be extracted from each tree, and also severely limits the total amount of TAXOL.RTM. which may be extracted, since new trees must be planted and matured before they can be stripped of their bark. The fact that yew trees grow very slowly further hinders attempts to produce TAXOL.RTM.. From an environmental standpoint, the constant uprooting of yew trees may have severe effects on the natural habitats that accommodate the trees.
Methods of extracting TAXOL.RTM. from yew tree leaves were reported in Korean Patent Nos. 95-10903, 95-25034 and 97-1339. These-patents pertain to techniques used to analyze the TAXOL.RTM. content in yew tree leaves. They do not disclose commercially feasible methods of the extraction of TAXOL from yew tree leaves. The primary element which determines the economic feasibility of the production of TAXOL.RTM. is the removal of chlorophyll from the yew tree leaves. Chlorophyll deactivates silica gel packing material, which is used in purification columns and is necessary for the purification of the extracts from the yew tree leaves.
The present invention provides a method that not only discloses the effective removal of chlorophyll from yew tree leaves, but also the extraction of substantially pure TAXOL.RTM. from the leaves. Specifically, the present invention discloses a process which yields 80-100 mg of at least 98% pure TAXOL.RTM. from 1 kg of yew tree leaves.